Bacillus velezensis bvx001, compound microbial agent and application

Abstract

The invention belongs to the technical field of microorganisms, and particularly relates to bacillus velezensis bvx001, a compound microbial agent and application. The bacillus velezensis bvx001 is preserved in China General Microbiological Culture Collection Center (CGMCC) on January 3, 2025, the address of the bacillus velezensis bvx001 is No.3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the preservation number of the bacillus velezensis bvx001 is CGMCC No.33291. The invention provides a compound microbial agent which has the capabilities of dissolving silicon, dissolving phosphorus and fixing nitrogen and has comprehensive functions. The soil conditioner can improve the content of silicon, phosphorus and nitrogen elements in soil, enhance plant stress resistance and promote plant growth, and can be used for development and utilization of saline-alkali soil.

Description

Technical Field

[0001] The present application belongs to the field of microbial technology, and specifically relates to a Bacillus velezinii bvx001, a composite microbial agent and its application. Background Art

[0002] Saline-alkali soil contains a large amount of salt ions, which makes the soil moisture content low and the air permeability poor; high concentration of salt ions destroys the osmotic balance and water balance of plants, reduces the photosynthetic rate of plants and limits the absorption of water by plants, resulting in difficulty in plant germination and loss of yield. In addition, the soil contains a large amount of nutrients, but the low solubility of nutrients caused by saline-alkali stress greatly hinders the absorption and utilization of plants, mainly including silicon, phosphorus, nitrogen and other elements, which play an important role in the growth and development of plants. Among them, nitrogen is a component of important biological macromolecules such as proteins and nucleic acids, which directly affects the growth, development and metabolism of crops; phosphorus plays an irreplaceable role in key physiological processes such as energy transfer and photosynthesis; and silicon can not only promote the formation and development of plant organs, but also play a more important role in improving the resistance of plants to biological and abiotic stresses. At present, many studies have shown that silicon can improve the resistance of plants in adversity and promote plant growth. Therefore, improving the bioavailability of silicon by plants in the soil has great research significance for the development and utilization of saline-alkali land. However, most silicon in the soil cannot be directly used by plants. The most common method is to apply additional silicon fertilizer. However, silicon fertilizer is easy to be lost after being applied to the soil and has a slow fertilizer effect, resulting in low utilization rate. In addition, due to the small amount of silicon fertilizer and the lack of unified standards, the quality of silicon fertilizer on the market is mixed. At the same time, most silicon fertilizers are expensive, which leads to high economic costs and little benefits. Therefore, a new solution is urgently needed. Summary of the invention

[0003] The purpose of the present invention is to solve the deficiencies of the prior art and provide a Bacillus velez bvx001, a composite microbial agent and application, which specifically adopts the following technical solutions: In a first aspect, the present invention provides a strain of Bacillus Velezii bvx001, wherein the Bacillus Velezii ( Bacillus velezensis ) bvx001 was deposited on January 3, 2025 at the General Microbiology Center of China Culture Collection Administration, located at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No.33291.

[0004] The gene sequence of 16S rDNA of the above-mentioned Bacillus velez bvx001 is shown in SEQ ID No.1.

[0005] SEQ ID No.1:

[0006] The silicon-dissolving bacteria provided by the present invention can convert insoluble silicates into silicon (i.e., monosilicic acid) that can be absorbed by plants through a series of biochemical reactions. In addition, the bacteria can colonize around the roots of the soil, form a symbiotic relationship with the plants, and play a sustainable role, thereby alleviating the damage of salt to the plants and promoting plant growth.

[0007] In a second aspect, the present invention provides a composite microbial agent, which includes the composite microbial agent mentioned above.

[0008] As a further preferred embodiment, the composite microbial agent further comprises Acinetobacter acin003; the Acinetobacter ( Acinetobacter sp. )acin003 was deposited on January 3, 2025 at the General Microbiology Center of China Culture Collection Administration, located at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No. 33292.

[0009] The gene sequence of 16S rDNA of the above-mentioned Acinetobacter acin003 is shown in SEQ ID No.2.

[0010] SEQ ID No.2:

[0011] As a further preferred embodiment, the composite microbial agent further comprises Exiguobacterium exio007; the Exiguobacterium ( Exiguobacterium sp. ) exio007 was deposited on January 3, 2025 at the General Microbiology Center of China Culture Collection Administration, located at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No. 33293.

[0012] The gene sequence of 16S rDNA of the above-mentioned Exiguobacterium exio007 is shown in SEQ ID No.3.

[0013] SEQ ID No.3:

[0014] The invention provides a composite microbial agent, wherein three strains of bacteria in the composite microbial agent all have certain salt and alkali resistance and growth-promoting abilities, and the composite agent simultaneously has silicon-solubilizing, phosphorus-solubilizing and nitrogen-fixing abilities, and according to a saline-alkali soil experiment of the invention, the strains are compounded into a BAE agent and inoculated into cabbage to study its influence on the growth of the cabbage, and the results show that compared with a single strain, the composite agent BAE can more significantly increase the silicon content in cabbage leaves, increase the leaf area, fresh weight and chlorophyll content of the cabbage, and improve the growth condition of the cabbage in a salt environment, and has a high practical application value.

[0015] In a third aspect, the present invention provides the use of the above-mentioned Bacillus Velezii bvx001 or the above-mentioned composite microbial agent in a product for promoting plant growth, wherein the product has the functions of simultaneously dissolving silicon, dissolving phosphorus and fixing nitrogen.

[0016] In a fourth aspect, the present invention provides the use of the above-mentioned Bacillus Velezii bvx001 or the above-mentioned composite microbial agent in promoting cabbage seed germination and / or preparing a cabbage seed germination promoter.

[0017] In a fifth aspect, the present invention provides a cabbage seed germination promoter, which comprises the above-mentioned Bacillus velezensis bvx001 or the above-mentioned composite microbial agent.

[0018] In a sixth aspect, the present invention provides the use of the above-mentioned Bacillus velezensis bvx001 or the above-mentioned composite microbial agent in inhibiting plant pathogens and / or preparing plant pathogen inhibitors.

[0019] As a further preferred embodiment, the plant pathogens include Fusarium graminearum var. avenae, Fusarium graminearum var. graminearum, Fusarium genus, Fusarium oxysporum, Fusarium graminearum, and Fusarium moniliforme.

[0020] In a seventh aspect, the present invention provides a plant pathogen inhibitor, which includes the above-mentioned Bacillus velezensis bvx001 or the above-mentioned composite microbial agent.

[0021] The beneficial effects of the present invention are: The present invention provides three salt-alkali tolerant bacterial strains with excellent silicon-dissolving ability, which have certain plant growth-promoting ability, can secrete auxin, iron carriers, etc., and can promote crop growth when applied in saline-alkali land. A composite microbial agent comprising the three strains of bacteria and several salt-alkali tolerant silicon-dissolving bacteria is also provided. The composite microbial agent has the ability to dissolve silicon, dissolve phosphorus, and fix nitrogen, and has comprehensive functions. It can increase the content of silicon, phosphorus, and nitrogen in the soil, enhance plant stress resistance and promote plant growth, and can be used for the development and utilization of saline-alkali land. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings required for use in the embodiments are briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without creative work.

[0023] Figure 1 Shown are the colony morphology and Gram staining of Bacillus velez bvx001, Acinetobacter acin003, and Exiguobacterium exio007 on LB plates, with the scale bar being 1 μm; Figure 2 Shown are the phylogenetic trees of Bacillus velez bvx001, Acinetobacter acin003, and Exiguobacterium exio007; Figure 3 Shown are the hydrolysis circles of Bacillus velez bvx001, Acinetobacter acin003, and Exiguobacterium exio007 on the hydrolysis plate; Figure 4 The figure shows the determination of silicon-degrading ability of Bacillus velez bvx001, Acinetobacter acin003, and Exiguobacterium exio007 in liquid; Figure 5 The figure shows the growth of Bacillus velez bvx001, Acinetobacter acin003, and Exiguobacterium exio007 in LB liquid medium supplemented with different concentrations of NaCl; Figure 6 Shown are the growth conditions of Bacillus velez bvx001, Acinetobacter acin003, and Exiguobacterium exio007 in LB liquid culture medium at different pH values; Figure 7 Shown are the phosphate hydrolysis circles of Acinetobacter acin003 and Exiguobacterium exio007; Figure 8 Shown is the nitrogen-fixing hydrolysis circle of Acinetobacter acin003; Fig. 9 Shown are the sizes of silicon hydrolysis circles of Bacillus velez bvx001, Acinetobacter acin003, and Exiguobacterium exio007 under different salt conditions; Fig.10 Shown is the effect of Bacillus velez bvx001, Acinetobacter acin003, and Exiguobacterium exio007 on the growth of Chinese cabbage under salt conditions; Fig.11 The figure shows the effect of Bacillus velez bvx001, Acinetobacter acin003 and Exiguobacterium exio007 on the physiological parameters of cabbage under salt conditions; Fig.12 The figure shows the antagonistic effect of Bacillus velezensis bvx001 on pathogenic fungi. A in the figure is a gramineous husk avena variety ( Gaeumonnomyces graminis var avenae ); B in the figure is a variety of Gramineae graminea ( Gaeumonnomyces graminis ); C in the figure is yellow Fusarium ( Fusariumgraminearum culmorum ); D in the figure is Fusarium oxysporum ( Fusariumgraminearum oxysporum ); E in the figure is Fusarium graminearum ( Fusariumgraminearum ); F in the figure is Fusarium moniliforme ( Fusarium verticillioides ). DETAILED DESCRIPTION

[0024] The following will be combined with the drawings in the embodiments of the present application to clearly and completely describe the technical solutions in the embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present application, not all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of this application.

[0025] Example 1 Screening of Bacillus velez bvx001, Acinetobacter acin003, and Exiguobacterium exio007 1. Prepare soil suspension: weigh 10.0 g of soil sample, add it to a sterilized 250 mL shake flask containing 90 mL of sterile water, and shake it in a shaker (200 rpm, 30 °C, 2 h~3 h) to obtain 10 -1 g / mL soil suspension; the soil suspension was diluted to 10 by 10-fold dilution method. -2 g / mL, 10 -3 g / mL, 10 -4 g / mL, 10 -5 g / mL, 10 -6 g / mL; 2. Purify the strain: Take 10 -4 g / mL, 10 -5 g / mL, 10 -6 100 μL of the 200 μg / mL dilution gradient suspension was evenly spread on Alexandrov solid medium (sucrose 5 g / L, Na2HPO4 2 g / L, MgSO4·7H2O 0.5 g / L, FeCl3·6H2O 0.005 g / L, CaCO3 0.1 g / L, magnesium silicate 2.5 g / L, agar 20 g / L) plates and placed in a 30 ℃ constant temperature incubator for 2d~3d. Several single colonies with characteristics related to silicon-dissolving microorganisms were picked and purified and streaked on Alexandrov solid plates, and repeated 2~3 times.

[0026] 3. Identification of strains: (1) Strain morphology observation and Gram staining The individual strains were screened out by the dilution spread plate method and named as: bvx001, acin003, exio007, streaked on LB solid plates (NaCl 10 g / L, peptone 10 g / L, yeast powder 5 g / L, agar 1.5 g / L), and cultured in an incubator at 30°C for 24 h, and the colony morphology was observed. Inoculate it into liquid LB medium (NaCl 10 g / L, peptone 10 g / L, yeast powder 5 g / L), take 10 μL of bacterial solution and drop it on a sterile glass slide, slightly heat the strain over the flame of an alcohol lamp to fix it, and after the glass slide cools down, add crystal violet solution until the colonies are covered, and stain for about 1 min; gently rinse off the stain with deionized water and wipe dry with absorbent paper; add iodine solution until the colonies are covered, and stain for about 1 min; rinse the stain in the same way and wipe dry; add 95% anhydrous ethanol to decolorize until the colonies are colorless, and wash with water; add tomato red stain until the colonies are covered, stain for about 1 min, rinse and dry in the same way, and then observe under a microscope.

[0027] The results are as follows Figure 1 As shown: The single colony of strain bvx001 on the LB solid plate is milky white, smooth, spherical, sticky and has neat edges. Under the microscope, the bacteria are rod-shaped and the Gram staining result is blue-purple, which is a Gram-positive bacterium.

[0028] The colonies of strain acin003 on the LB solid plate are grayish white, spherical, with smooth surfaces and neat edges. Under a microscope, the bacteria are coccobacilli, often arranged in pairs. The Gram staining result is purple-red, and it is a Gram-negative bacterium.

[0029] The colonies of strain exio007 on the LB solid plate are orange-yellow, round, and moist on the surface. Under a microscope, the bacteria are short rods, arranged singly or in pairs. The Gram staining result is blue-purple, and it is a Gram-positive bacterium.

[0030] (2) Molecular biological identification The genomes of the three strains were extracted using a bacterial DNA extraction kit and used as a template. The 16SrDNA universal primer sequences 27F (SEQ ID No.4) 5'-AGAGTTTGATCCTGGCTCAG-3' and 1492R (SEQ ID No.5) 5'-GGTTACCTTGTTACGACTT-3' were used to amplify the screened strains by PCR. After completion, the strains were sent to Shanghai Jie Li Biotechnology Co., Ltd. for sequencing. The sequencing results are shown in SEQ ID No.1, SEQ ID No.2, and SEQ ID No.3.

[0031] Table 1 PCR reaction system The obtained sequences were submitted to NCBI for Blast sequence alignment, and the evolutionary tree was constructed using mega11 software (Molecular Evolutionary Genetics Analysis).

[0032] The results show that: Phylogenetic tree of the three strains Figure 2 As shown, bvx001 was identified as Bacillus velezinsis; acin003 was identified as Acinetobacter; and exio007 was identified as Exiguobacterium.

[0033] Example 2 Detection of silicon decomposition ability of Bacillus velez bvx001, Acinetobacter acin003 and Exiguobacterium exio007 (1) Flat panel legal testing Take 10 μL of bacterial solution and place it on magnesium silicate plates (glucose 10 g / L, magnesium silicate 2.5 g / L, agar 1.5 g / L) to observe the size of the hydrolysis circle. The results are as follows: Figure 3 As shown: the three strains of bacteria can form silicon-dissolving hydrolysis circles on the magnesium silicate plate, indicating that these three strains of bacteria all have the ability to dissolve silicon.

[0034] (2) Liquid method quantitative detection The cells were inoculated into Alexander Rof liquid medium at a 1% inoculation rate and cultured at 30°C and 200 rpm for 5 days. The culture solution was centrifuged at 8000 rpm for 15 min, the supernatant was filtered, and the soluble silicon content in the supernatant was detected by inductively coupled plasma-mass spectrometry.

[0035] The experimental results are as follows Figure 4 As shown: Compared with the blank control CK without added bacteria, the soluble silicon content in the culture medium increased to a certain extent after inoculation with the three strains of bvx001, exio007 and acin003. Among them, the change in the group with added bvx001 strain was the most significant, followed by exio007 and acin003.

[0036] Example 3 Detection of salt-alkali resistance and growth-promoting ability of Bacillus velez bvx001, Acinetobacter acin003 and Exiguobacterium exio007 (1) Salt and alkali resistance Single colonies were picked and grown in LB liquid culture medium overnight, and inoculated at a 1% inoculum size into liquid LB containing 1% NaCl (w / v), 3% NaCl (w / v), 5% NaCl (w / v), 6% NaCl (w / v), 7% NaCl (w / v), 8% NaCl (w / v), 9% NaCl (w / v), 10% NaCl (w / v), and 11% NaCl (w / v) and liquid LB culture medium with pH = 7, 8, 9, 10, and 11, respectively. The cultures were cultured at 30°C, 200 rpm constant temperature shaking incubator for 24 h, and the OD600nm of the bacterial solution was determined by a spectrophotometer.

[0037] The results are as follows Figure 5 As shown: the strain with the strongest salt tolerance is exio007, when the NaCl concentration is 7% (w / v), about 50% of the strains can grow and reproduce; the second is strain bvx001, compared with 1% NaCl (w / v), under 7% NaCl (w / v) conditions, about 30% of the strains can survive; finally, acin003, although not as salt-tolerant as the other two strains, it also has a certain salt tolerance and can tolerate up to 6% NaCl (w / v).

[0038] The results are as follows Figure 6 As shown: the three strains have strong alkali resistance, among which acin003 and exio007 can tolerate an alkaline environment of pH=10 at most, and bvx001 can tolerate an alkaline environment of pH=9 at most.

[0039] (2) Determination method of auxin (IAA) Single colonies were picked and grown overnight in LB liquid medium; 1% inoculation was inoculated into LB liquid medium containing L-tryptophan (200 mg / L), and cultured in a shaking incubator at 180 r / min and 30°C for 36 h; 2 mL of bacterial solution was pipetted into a centrifuge tube, centrifuged at 10000 r / min for 10 min, 1.5 mL of supernatant was taken and added with an equal volume of Salkowski colorimetric solution (50 mL 35% HClO4 + 1 mL 0.5 mol / L FeCl3) in a 5 mL centrifuge tube, and kept in the dark for 30 min, and its OD530 nm value was determined. The standard solution was made with analytically pure indoleacetic acid and then subjected to Salkowski colorimetric reaction to obtain a standard curve. The content of IAA per unit volume of fermentation broth was calculated by comparing with the standard curve.

[0040] The results are shown in Table 2. The results showed that all three strains could secrete auxin, among which bvx001 had the highest production, up to 54.4±0.22 mg / L, compared with the negative control Escherichia coli DH5-α, which was only 8.0 mg / L.

[0041] Table 2 The production of longin by different strains (3) Determination method of siderophore Pick a single colony and grow it in LB liquid culture medium overnight; inoculate it in MKB (full name) liquid culture medium (acid hydrolyzed casein 5 g / L, MgSO4·7H2O 2.5 g / L, glycerol 15 mL / L, K2HPO4·3H2O 3.28 g / L) at a 1% inoculation volume and culture it in a 30 ℃ shaking constant temperature incubator for 36 h; take 1 mL of bacterial solution and centrifuge it at 12000 rpm for 10 min, take 100 μL of supernatant and mix it with 100 μL of chrome azure (CAS) quantitative detection solution, use sterile ddH2O for the same operation as a control, react at room temperature for 1 hour, record the OD630nm values ​​of the sample (As) and the control (Ar), and calculate the iron carrier yield using the following formula: Siderophore unit (%) = 100 × (Ar—As) / Ar The results are shown in Table 3. The results show that all three strains can secrete iron carriers, indicating that they all have certain growth-promoting abilities. The iron carrier secreted by bvx001 is as high as 87%, and that of exio007 is as high as 73%, which are strains with high efficiency in secreting iron carriers.

[0042] Table 3 Siderophore secretion of different strains (4) Determination of ACC deaminase production capacity a. Pick a single colony and place it in 30 mL of DF liquid medium (KH2PO4 4 g / L, Na2HPO4 6 g / L, MgSO4·7H2O 0.2 g / L, glucose 2 g / L, gluconic acid 2 g / L, citric acid 2 g / L, H3BO3 0.01 mg / L, MnSO4·H2O 0.01 mg / L, ZnSO4·7H2O 0.078 mg / L, CuSO4·5H2O 0.05 mg / L, MoO3 0.01 mg / L, FeSO4·7H2O 1 mg / L), and culture it in the dark at 30 ℃ and 200 r / min with shaking; b. Centrifuge the bacterial suspension at 4°C and 8000 r / min for 10 min, collect the bacterial suspension, and resuspend the precipitate in 30 mL of ADF (DF medium supplemented with 5 mmol / L ACC solution) liquid medium, and culture at 30°C and 200 r / min in the dark for 24 h. c. Centrifuge again at 4 °C and 8,000 r / min for 10 min, resuspend with 5 mL of 0.1 mol / L Tris-HCl buffer (pH 7.6), wash twice; transfer to a 1.5 mL centrifuge tube, resuspend the cells in 600 μL of 0.1 mol / L Tris-HCl buffer (pH=8.5) to dissolve the cells; add 30 μL of toluene and vortex for 30 s to break the cells; d. Enzyme activity determination: Pipette 200 μL of broken cell suspension, add 20 μL of 0.5 mol / L ACC solution, mix well and heat in a water bath at 30 ℃ for 15 min; e. Then add 1 mL of 0.56 mol / L HCl solution, shake well, and centrifuge at 10000 r / min for 10 min; f. Take 1 mL of the above centrifugal supernatant, add 800 μL of 0.56 mol / L HCl solution, mix well, then add 300 μL of 2 g / L 2, 4-dinitrophenylhydrazine solution, and incubate in a 30 ℃ water bath for 30 min; g. Add 2 mL of 2 mol / L NaOH solution for color development and detect the OD540 value.

[0043] After deducting the spontaneous product in the control sample, the OD540 value of each strain was substituted into the standard curve to calculate the corresponding α-ketobutyric acid content, and the amount of 1 μmoL α-ketobutyric acid formed per minute was defined as 1 enzyme activity unit.

[0044] (6) Determination of cell protein concentration Take 10 μL of toluene-treated bacterial cells and dilute them 10 times with 1×PBS; dilute 1 mg / mL BSA protein standard solution with 1×PBS to obtain 20 μg mL -1 , 40 μg·mL -1 , 60 μg·mL -1 , 80 μg·mL -1 , 100 μg·mL -1 The protein concentration gradient was plotted, with 1×PBS as blank control, and each concentration was repeated 3 times; the protein concentration was measured using a Bradford protein kit, and the OD595 value of each well was measured using an ELISA instrument after 5 minutes of reaction. The standard curve was drawn with the average OD595 value as the ordinate and the corresponding protein concentration as the abscissa; the protein concentration of the sample dilution was determined on the standard curve based on the average OD595 value of the sample dilution.

[0045] The results are shown in Table 4: Bacillus velez bvx001 can produce ACC deaminase, and the enzyme activity produced can reach 2.733 ± 0.55 U / mg. Escherichia coli DH5-α is used as a negative control, and the enzyme activity produced by Escherichia coli DH5-α is 0.0073 U / mg.

[0046] Table 4 ACC deaminase activity of different strains (5) Determination of phosphorus solubilizing capacity Take 10 μL of bacterial solution and culture it in Montgena inorganic phosphate medium (PKO) (glucose 10 g / L, (NH4)2SO4 0.5 g / L, KCl 0.3 g / L, NaCl 0.3 g / L, FeSO4· 7H2O 0.03 g / L, MgSO4·7H2O 0.3 g / L, MnSO4· 4H2O 0.03 g / L, Ca3(PO4)2 10 g / L, agar 15 g / L) in an incubator at 30 ℃ for 5 d~7 d. If a transparent hydrolysis circle is produced, it means that it has the ability to solubilize phosphorus.

[0047] The results are as follows Figure 7 As shown: On the PKO plate, transparent hydrolysis zones appeared around the colonies of acin003 and exio007, indicating that the strains also have the ability to solubilize phosphate.

[0048] (6) Determination of nitrogen fixation capacity Take 10 μL of bacterial solution and place it on an Ashby nitrogen-free medium plate (KH2PO4 0.2 g / L, NaCl 0.2 g / L, MgSO4 0.2 g / L, CaCO3 5 g / L, CaSO4 0.1 g / L, glucose 10 g / L, agar powder 20 g / L) and culture it in a 30 ℃ incubator for 5 to 7 days. If a transparent hydrolysis circle is produced, it means that the bacteria has the ability to fix nitrogen.

[0049] The results are as follows Figure 8 As shown: On the Ashby nitrogen-free medium plate, a transparent hydrolysis zone appeared around the acin003 colony, indicating that the strain has the ability to fix nitrogen.

[0050] Example 4 Determination of silicon-degrading ability of Bacillus velez bvx001, Acinetobacter acin003 and Exiguobacterium exio007 under salt conditions 10 μL of bacterial solution was placed on magnesium silicate plates with different NaCl concentrations. The NaCl concentration was set to 3% NaCl (w / v) and 6% NaCl (w / v), and the size of the hydrolysis circle was then observed and recorded.

[0051] The experimental results are as follows Fig. 9 As shown: On the two plates with additional NaCl, the three strains still have the ability to degrade silicon, and under the condition of 3% NaCl (w / v) medium, the silicon degrading efficiency (D / d) of aci003 and exio007 is higher than that under normal conditions, but as the salt concentration continues to increase, the hydrolysis circle formed by the strains' silicon degrading shrinks again, indicating that the high-salt environment will inhibit the strains' silicon degrading ability. In addition, the three strains still have the ability to degrade silicon on the plates with added NaCl, which shows that the composite bacterial agent has potential application value in the improvement of saline-alkali land.

[0052] Table 5 Silicon-degrading ability of strains at different salinities Example 5 To explore the growth-promoting effect of a composite microbial agent consisting of Bacillus velez bvx001, Acinetobacter acin003 and Exiguobacterium exio007 on Chinese cabbage The pots used in the experiment had a capacity of about 5.5 L, and each pot contained 2.0 kg of soil. The soil was irrigated with different concentrations of sodium chloride solution (3.0 g / kg and 4.5 g / kg) in advance and stirred evenly. It was placed for 1 to 2 days to make it closest to the natural state. Cabbage seeds with full grains were selected and sown in a triangular pattern, with 3 seeds per pot. The bacterial suspension was inoculated into the soil around the cabbage seeds in each pot at a 10% inoculation rate. The blank control group was replaced with an equal amount of clean water and placed in a natural environment for growth. After that, tap water was used for irrigation, and insoluble magnesium silicate was added to the water for irrigation, with an additional amount of 2 g / kg. Harvest after about 50 days.

[0053] Table 6 Potted plant experimental groups It should be noted that in the above table, BE represents a composite microbial agent of Bacillus belezii bvx001 and Exiguobacterium exio007; AE represents a composite microbial agent of Acinetobacter acin003 and Exiguobacterium exio007; AB represents a composite microbial agent of Bacillus belezii bvx001 and Acinetobacter acin003; BAE represents a composite microbial agent of Bacillus belezii bvx001, Acinetobacter acin003 and Exiguobacterium exio007.

[0054] After 50 days of cultivation, the physiological conditions of each group of cabbage plants were recorded, and their physical indicators were measured: root weight and leaf area, etc.; biochemical indicators: silicon content, total chlorophyll and malondialdehyde content in leaves.

[0055] The experimental results are as follows Figure 10-11As shown in the pot experiment, salt stress can significantly inhibit the normal growth and development of plants, resulting in dwarfing of plants, withering and yellowing of leaves, etc. After adding different bacterial suspensions under salt stress, the growth state of cabbage is significantly better than that of plants treated with simple salt stress, and the same salt-tolerant strains have different mitigation effects. For example, after adding only the bvx001 strain, the leaf area of ​​cabbage under the two salt gradients increased by 68% and 46%, the root weight increased by 61% and 120%, and the fresh weight increased by 30% and 65%, respectively, which also has a relatively obvious growth-promoting effect. At the same time, it can be observed that the effect of adding the composite microbial agent was the most significant. Under the 3.0 g / kg and 4.5 g / kg NaCl environments, the leaf area increased by 173% and 186% respectively; the fresh weight increased by 140% and 300% respectively, even higher than that of the cabbage in the blank control group; at the same time, under the condition of 4.5 g / kg NaCl, the root weight of cabbage was most obviously inhibited by salt stress, which was significantly lower than that of the blank control, while the root weight was significantly increased after inoculation with the composite microbial agent, with an increase of 200%.

[0056] By measuring the changes in silicon content in leaves, it was found that salt stress affected the plant's absorption of nutrients including silicon in the soil, which led to yellowing and dwarfing of the leaves, resulting in lower silicon content in the leaves. Under the condition of additional addition of insoluble magnesium silicate to each pot, the silicon content in the cabbage leaves inoculated with silicon-removing bacteria increased to a certain extent compared with the salt control group, especially the cabbage inoculated with the composite bacterial agent group, which reached a significant difference compared with the control group. In addition, salt stress inhibits the photosynthesis of plants, resulting in a decrease in the total chlorophyll content of plants. Compared with CK, the content of photosynthetic pigments in the cabbage plants under salt stress treatment decreased. Under the treatment of 3.0 g / kg salt, the total chlorophyll content increased by 40% and 137% after adding a single strain bvx001 and a composite bacterial agent, respectively; when the salt concentration was increased to 4.5 g / kg, the total chlorophyll content of the two groups increased by 17% and 67%, respectively. In addition, the strain can also reduce the content of malondialdehyde (MDA) in plants, which is a commonly used indicator. When plant organs age or suffer damage under adverse conditions, membrane lipid peroxidation often occurs. MDA is the final decomposition product of membrane lipid peroxidation, and its content can reflect the degree of damage suffered by the plant under adverse conditions. After inoculation with bvx001, MDA decreased by 8% and 34% under the two salt treatments, respectively; when the BAE composite bacterial agent was inoculated into the environment, MDA decreased by 58% and 68% under the two salt treatment conditions, respectively, which was a significant difference. In summary, the composite bacterial agent BAE in this patent significantly promotes the growth and development process of cabbage under salt stress, and can effectively alleviate the physiological damage caused by salt to cabbage, thereby enhancing the stress resistance of cabbage, showing a high application value in salinized soil improvement and salt stress agricultural production practice.

[0057] Example 6 Experiment on inhibition of pathogens by Bacillus velez bvx001, Acinetobacter acin003 and Exiguobacterium exio007 Use a sterile toothpick to pick up a fungal block of about 2 cm × 2 cm and invert it in the center of the PDA solid plate. Add equal amounts of bacterial solution with the same OD to the three corners of the plate. There are three strains in the experiment: bvx001, acin003 and exio007. After culturing in a 26°C incubator for 5 days, the antibacterial results were observed. The pathogens are shown in Table 7.

[0058] Table 7 List of pathogens The experimental results showed that among the 8 strains tested, only bvx001 had an inhibitory effect on pathogenic fungi. Fig.12 As shown, bvx001 has a significant inhibitory effect on 6 of the strains. Based on the antibacterial results of bvx001, the strain can mainly inhibit the fungi of the genus Fusarium, which mainly cause plant root rot, stem rot, stem base rot, etc., such as wheat fusarium, wheat rust and corn stalk rot. It can be seen that the strain bvx001 has potential biological control ability. It is applied to the preparation of composite microbial agents. On the one hand, it can promote the growth of plants under salt stress environment, and on the other hand, it can effectively inhibit the growth and spread of specific pathogens, which has a high application value in the field of agricultural production.

[0059] The embodiments of the present application are described above in conjunction with the accompanying drawings. Specific examples are used herein to illustrate the principles and implementation methods of the present application. The description of the above embodiments is only used to help understand the core idea of ​​the present application, but the present application is not limited to the above-mentioned specific implementation methods. The above-mentioned specific implementation methods are merely illustrative and not restrictive. Under the inspiration of the present application, ordinary technicians in this field can also make many forms without departing from the scope of protection of the purpose of the present application and the claims, all of which are within the protection of the present application.

Claims

1. A strain of Bacillus velez bvx001, characterized in that The Bacillus Velez Bacillus velezensis ) bvx001 was deposited on January 3, 2025 at the General Microbiology Center of China Culture Collection Administration, located at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No.33291.

2. A composite microbial agent, characterized in that: The composite microbial agent includes the Bacillus Velezii bvx001 described in claim 1.

3. The composite microbial agent according to claim 2, characterized in that: The composite microbial agent also includes Acinetobacter acin003; the Acinetobacter ( Acinetobacter sp. )acin003 was deposited on January 3, 2025 at the General Microbiology Center of China Microorganism Culture Collection Administration, located at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No.33292.

4. The composite microbial agent according to any one of claims 2 to 3, characterized in that: The composite microbial agent also includes Exiguobacterium exio007; the Exiguobacterium ( Exiguobacterium sp. ) exio007 was deposited on January 3, 2025 at the General Microbiology Center of China Microorganism Culture Collection Administration, located at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No.33293.

5. Use of the Bacillus Velezii bvx001 according to claim 1 or the composite microbial agent according to any one of claims 2 to 4 in a product for promoting plant growth, characterized in that: The product has the functions of dissolving silicon, dissolving phosphorus and fixing nitrogen.

6. Use of the Bacillus velez bvx001 described in claim 1 or the composite microbial agent described in any one of claims 2 to 4 in promoting cabbage seed germination and / or preparing a cabbage seed germination promoter.

7. A cabbage seed germination promoter, characterized in that The cabbage seed germination promoter includes the Bacillus velezensis bvx001 described in claim 1 or the composite microbial agent described in any one of claims 2-4.

8. Use of the Bacillus velezensis bvx001 described in claim 1 or the composite microbial agent described in any one of claims 2 to 4 in inhibiting plant pathogens and / or preparing plant pathogen inhibitors.

9. The use according to claim 8, characterized in that: The plant pathogens include Fusarium graminearum var. avena, Fusarium graminearum var. graminearum, Fusarium oxysporum, Fusarium graminearum and Fusarium moniliforme.

10. A plant pathogen inhibitor, characterized in that: The plant pathogen inhibitor includes the Bacillus velezensis bvx001 described in claim 1 or the composite microbial agent described in any one of claims 2-4.

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